Using limonen-10-ol to kill bacteria, yeast, and fungi

ABSTRACT

Bacteria, fungi, or yeast are treated in their habitat with lethal concentrations of limonen-10-ol.

TECHNICAL FIELD

The object of this invention is to demonstrate a method of using LIMONEN-10-OL to kill bacteria, yeast, and fungi.

BACKGROUND OF THE INVENTION

(1) Field of the invention

During the study of limonene as a hand cleaner, it was found that limonene can be made bactericidal and fungicidal as was illustrated by the applicants in U.S. Pat. Nos. 5,153,229 and 5,229,425. A review of the literature showed that oxidized limonene contains several oxidation products including cis and trans-carveol, trans-p-menth-8-ene-1,2-diol, limonene 1,2-epoxide, limonene 8,9-epoxide, cis and trans-p-mentha-2,8-dien-1-ol, and perillyl alcohol, as was outlined by Blumann in Chemical Abstracts, Volume 63, 1965, on page 1819. The applicants decided to study other monocyclic monoterpenes for antimicrobial activity. Because limonene is not bactericidal and because the chemical structures of limonene and limonen-10-ol are identical except for a hydroxyl group replacing a hydrogen atom at carbon 10, the applicants were surprised to find that limonen-10-ol is bactericidal and fungicidal. A review showed that all the monocyclic monoterpenes that are known to be antimicrobial have an oxygen molecule or hydroxyl group replacing a hydrogen atom at carbons 2, 3, 4, or 8. Prior to the discovery by the applicants that perillyl alcohol is bactericidal, as demonstrated in U.S. Pat. No. 5,110,832, and menth-1-en-9-ol is bactericidal and fungicidal as outlined in U.S. Pat. No. 5,294,645 no monocyclic monoterpene with an oxygen or a hydroxyl group at carbon 7, 9 or 10 was recognized as having antimicrobial activity. After the applicants discovered that perillyl alcohol is bactericidal, the applicants decided to study limonen-10-ol for antimicrobial activity and were pleasantly surprised to find that limonen-10-ol kills bacteria, yeast, and fungi in bactericidal and fungicidal concentrations.

The chemical structures of limonene and limonen-10-ol follow below. ##STR1##

Because limonene is not bactericidal nor fungicidal and because the chemical structures of limonene and limonen-10-ol are identical except for a hydroxyl group replacing a hydrogen atom at carbon 10, limonen-10-ol was not expected to be bactericidal and fungicidal. It should be noted in the structures of the monocyclic monoterpenes that follow, that none of the monocyclic monoterpenes that are known to be bactericidal, have an oxygen atom or a hydroxyl group at carbon 9 or 10 as can be appreciated in the structures of: perillyl alcohol, carveol, carvone, dihydrocarveol, dihydrocarvone pulegone, isopulegol, menthol, menthone, terpinen-4-ol, and a-terpineol which follow. ##STR2##

Limonen-10-ol is an oil with a fruity aroma that is soluble in alcohol and is miscible in oil. Although it is insoluble in water, it can be easily emulsified in water. It is poorly soluble in propylene glycol, and is almost insoluble in glycerine. Limonen-10-ol occurs naturally, but heretofore, it has not been used as a bactericide nor fungicide.

Limonen-10-ol is available commercially from the Aldrich Chemical Company, 940 Saint Paul Avenue, Milwaukee, Wis. 53233, catalogue number 21,841-3, and from George Majetich PhD, Department of Chemistry, University of Georgia, Athens, Ga. 30602. It is produced by a proprietary method of oxygenating d-limonene.

(2) Description of the Prior Art

Zuckerman studied the effect of auto-oxidized d-limonene on bacteria, and found it was weakly bacteriostatic, was unstable, and lost its bacteriostatic effect on keeping as was discussed in Nature 168:517 (1961). He never studied limonen-10-ol. Kurita investigated the fungicidal activity of several components of essential oils as was reported in Biol. Chem., 45(4), 945-952, 1981, but he never studied the antimicrobial activity of limonen-10-ol against bacteria, yeast nor fungi. Murdock and Allen showed the germicidal effect of sodium benzoate against yeast was enhanced by orange peel oil and d-limonene, as was outlined in Food Technology Vol 14, No 9, 1960, pages 441-5. They never studied the activity of limonen-10-ol against bacteria, yeast nor fungi. Kellner et al demonstrated that ethereal oils and some of their components have anti-bacterial activity as was reported in Arneimittel-Forechung, 5, 224-9, 1955. He confirmed that limonene is not bactericidal. He never studied limonen-10-ol for bactericidal nor fungicidal activity. Gauvreau showed a means of producing disinfecting compositions in U.S. Pat. No. 3,595,975 by combining cetyl pyridinium with terpenes to form antiseptics, but he never studied limonen-10-ol alone nor in combination with cetyl pyridinium hydrochloride. A. Morel revealed the sterilizing action of carveol, dihydrocarveol, and their ozonization products in Comp. Rend. Soc. Biol. Volume 115, pages 536-8 (1934). He demonstrated the bactericidal effect of carveol and dihydrocarveol, but he never studied the bactericidal nor fungicidal activity of limonen-10-ol. The applicants discovered that limonene can be made bactericidal and fungicidal by oxidation as was outlined in U.S, Pat. Nos. 5,153,229 and 5,229,425 but limonen-10-ol was never studied for bactericidal nor fungicidal activity and limonene-10-ol has never been identified in oxidized limonene.

It should be pointed out that drugs which are bactericidal are usually not fungicidal, and drugs which are fungicidal are usually not bactericidal. In addition, drugs which are bactericidal frequently promote the growth of yeast. Table A, which follows, exemplifies the bactericidal and fungicidal activity of several commonly used anti-bacterial, anti-yeast, and anti-fungal antibiotics.

                                      TABLE A                                      __________________________________________________________________________              ANTIBIOTIC ACTIVITY AGAINST                                           ANTIBIOTICS                                                                             Gm.sup.+ Bac                                                                          Gm.sup.- Bac                                                                          AFBact                                                                               Yeast                                                                               Fungi                                        __________________________________________________________________________     A. ANTIBACTERIAL                                                                1.                                                                              Ampicillin                                                                            YES    YES    NO    NO   NO                                            2.                                                                              Cephalothin                                                                           YES    YES    NO    NO   NO                                            3.                                                                              Chloram-                                                                              YES    YES    NO    NO   NO                                             phenicol                                                                      4.                                                                              Erythromycin                                                                          YES    NO     NO    NO   NO                                            5.                                                                              Ethambutol                                                                            NO     NO     YES   NO   NO                                            6.                                                                              Gentamicin                                                                            YES    YES    NO    NO   NO                                            7.                                                                              Isoniazid                                                                             NO     NO     YES   NO   NO                                            8.                                                                              Nitro- NO     YES    NO    NO   NO                                             furantoin                                                                     9.                                                                              Penicillin                                                                            YES    NO     NO    NO   NO                                           10.                                                                              Rifampin                                                                              YES    NO     YES   NO   NO                                             Streptomycin                                                                          YES    YES    YES   NO   NO                                             Sulfonamides                                                                          NO     YES    NO    NO   NO                                             Tetracycline                                                                          YES    YES    NO    NO   NO                                             Vancomycin                                                                            YES    YES    NO    NO   NO                                           B. ANTIYEAST                                                                    1.                                                                              Nystatin                                                                              NO     NO     NO    YES  NO                                            2.                                                                              Gentian                                                                               NO     NO     NO    YES  NO                                             violet                                                                       C. ANTIFUNGAL                                                                   1.                                                                              Chlotri-                                                                              NO     NO     NO    YES  YES                                            mazole                                                                        2.                                                                              Griseofulvin                                                                          NO     NO     NO    NO   YES                                          __________________________________________________________________________      Gm.sup.+ Bac = Gram Positive Bacteria, Gm.sup.- Bac = Gram Negative            Bacteria, AFBac = Acid Fast Bacteria, YES = Kills Organism, NO = No            Activity Against Organism                                                

It should be noted from the table above that none of the anti-bacterial antibiotics kill yeast nor fungi, and none of the anti-yeast nor anti-fungal antibiotics kill bacteria. Thus, an anti-fungal or anti-yeast antibiotic is not expected to kill bacteria and an anti-bacterial antibiotic is not expected to kill yeast nor fungi. Anti-fungal antibiotics do not necessarily kill yeast and anti-yeast antibiotics do not necessarily kill fungi.

DISCLOSURE OF THE INVENTION

This invention relates to the use of limonen-10-ol as an antimicrobial to kill bacteria, yeast, and fungi. Limonen-10-ol is an oil that is available commercially, but heretofore, it has not been recognized as an antimicrobial. It is slightly viscous and when applied, readily adheres to glass, metal, wood, cloth, rope, book covers, paper, paint, cement, ceramics, plastic, plant surfaces, skin, mucous membranes, and teeth leaving an oily film. Because it adheres to surfaces and is not soluble in water, it allows prolonged exposure and makes limonen-10-ol an ideal antimicrobial for treatment of infected plants, animals and humans, regardless of whether they are infected with bacteria, yeast, or fungi.

The exact method of killing bacteria, yeast, and fungi is unknown, but it is thought that limonen-10-ol kills bacteria, yeast, and fungi by lysing the cell membrane of the organisms which is lethal to the organisms.

In practice, any surface, on which it is desirable to kill or prevent the growth of bacteria, yeast or fungi, is treated with bactericidal, fungicidal, and/or effective concentrations of limonen-10-ol by swabbing, wiping, painting, washing, brushing, spraying, or any other direct application technique. Alternatively, limonen-10-ol can be incorporated in creams, ointments, tinctures, gels, suppositories, paints, sprays, sealers, aerosols, toothpastes, solutions, emulsions, soaps, scrubs, mouthwashes, or antiseptic, and applied anywhere it is desirable to kill or prevent the growth of bacteria, yeast, and fungi.

BEST MODE FOR CARRYING OUT THE INVENTION

The following examples are illustrative of the best mode of carrying out the invention. They are, obviously, not to be construed as limitative of the invention since various other embodiments can readily be evolved in view of the teachings provided herein.

EXAMPLE 1

ANTIMICROBIAL ACTIVITY OF LIMONEN 10-OL AGAINST BACTERIA, YEAST, AND FUNGI

The ANTIMICROBIAL contemplated by this invention is LIMONEN-10-OL which was studied for bactericidal, anti-yeast and fungicidal activity. The organisms tested included the bacteria: Staphylococcus aureus ATCC 25923, Streptococcus mutans which causes dental plaque, Escherichera coli 7, Salmonella 14 (para B), Pseudomonas aeriginosa 115, the yeast Candida albicans, a common cause of skin, mouth, and vaginal infections, and the fungi Cladosporium cladosporides, and Aueroblasidium pullans. The bactericidal concentration, minimal effective concentration, and the fungitidal concentration of limonen-10-ol needed to kill these organisms are outlined in Table B below. The limonen-10-ol used in these tests was obtained from George Majetich PhD, Department of Chemistry, University of Georgia, Athens, Ga. 30602, was 99.9% pure by gas chromatography, and its structure was verified by NMR mass spectroscopy. Limonen-10-ol can be obtained from the Aldrich Chemical Company 940 West Saint Paul Avenue, Milwaukee, Wis. 53201; its catalogue number is 21,841-3.

                  TABLE B                                                          ______________________________________                                         BACTERICIDAL AND FUNGICIDAL ACTIVITY                                           OF LIMONEN-10-OL                                                               ORGANISM          10 Min   60 Min   24 Hrs                                     ______________________________________                                                           MINIMAL BACTERICIDAL                                         A. BACTERIA       CONCENTRATION                                                1.  Staphylococcus aureus,                                                                           0.06     0.01   0.005                                        ATCC 25923                                                                 2.  Streptococcus mutans                                                                             0.02     0.01   0.005                                    3.  Escherichera coli 7                                                                              0.01     0.005  0.005                                    4.  Salmonella 14 (para B)                                                                           0.005    0.005  0.005                                    5.  Pseudomonas aeruginosa                                                                           0.3      0.1    0.06                                         ATCC 115                                                                   B. YEAST                                                                       1.  Candida albicans  0.01     0.02   0.005                                                      MINIMUM FUNGICIDAL                                           C. FUNGI          CONCENTRATION                                                1.  Cladosporium cladosporides                                                                       0.01     0.02   0.005                                        OM489                                                                      2.  Aureoblasidium pullans                                                                           0.005    0.0025 0.0025                                       OM279C                                                                     ______________________________________                                    

The standard assay used to test the bacterial activity of limonen-10-against the different strains of bacteria and yeast was as follows: various dilutions of limonen-10-ol were prepared in an appropriate broth medium for each test strain. An inoculum of 10⁶ colony-forming units (CFU)/ml was used. Each test was incubated at the proper temperature for each organism and subcultured (0.01 ml) at 10 minutes, 60 minutes, and 24 hours onto agar media free of limonen-10-ol. Results were expressed as the bactericidal concentration or the minimum effective concentration, i.e. the lowest concentration of limonen-10-ol (ml. limonen-10-ol/total ml of test) killing at least 99.99% of the bacterial or yeast inoculum.

The standard assay used to test the activity of limonen-10-ol against fungi was as follows: various dilutions of limonen-10-ol were prepared in Sabouraud dextrose broth medium. An inoculum of 10⁶ colony-forming units (CFU/ml) of fungi was introduced into each test, after which it was incubated at 37° C. in air, and subcultured (0.01 ml) at 10 minutes, 60 minutes, and 24 hours onto agar media free of 1imonen-10-ol. Results were expressed as the minimal fungicidal concentration, i.e. the lowest concentration of limonen-10-ol (ml limen-10-ol/total ml test), with no detectable viable colonies following subculture onto media free of limen-10-ol.

                                      TABLE C                                      __________________________________________________________________________     Test conditions used to assay the bactericidal activity of                     limonen-10-ol:                                                                                     SUBCULTURE                                                                               INCUBATION                                       ORGANISM  BROTH MEDIUM                                                                             AGAR MEDIUM                                                                              CONDITIONS                                       __________________________________________________________________________       Staphylococcus,                                                                        Mueller-  5% sheep  Air @                                              Enterobac-                                                                             Hinton    blood     37° C.                                      teriacea, and                                                                  Pseudomonas                                                                    Streptococcus                                                                          Todd-     5% sheep  10% CO.sub.2                                               Hewitt    blood     in air                                                                         at 37° C.                                   Yeast   Sabouraud 5% sheep  air at                                                     dextrose  blood     37° C.                                      Fungi   Sabouraud dextrose with                                                                            air at                                                     0.05% yeast extract 37° C.                                    __________________________________________________________________________

EXAMPLE 2

FORMULATIONS WHICH INCORPORATE LIMONEN-10-OL AS THE ANTIMICROBIAL COMPOUND TO KILL BACTERIA, YEAST, AND FUNGI

The following formulations are prepared using limonen-10-ol in liquids, gels, soaps, pastes, creams, ointments, suppositories, tampons, aerosols, paints, and emulsions. When bacteria, yeast, and fungi are treated with limonen-10-ol containing formulations, the formulations kill or prevent the growth of bacteria, yeast, and fungi.

A. LIQUIDS

1. Solutions or Sprays

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         a.   Limonen-   5.0%         0.1-50%                                                                               fungi-                                          10-ol                          cide                                            Corn Oil   95.0%       50-99.9%                                                                               diluent                                                   100.0%                                                          b.   Limonen-   1.0%         0.1-50%                                                                               bacteri-                                        10-ol                          cide                                            Ethyl      99.0%       50-99.9%                                                                               diluent                                         Alcohol   100.0%                                                          ______________________________________                                    

2. Mouthwash

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         a.   Limonen-   5.0%         0.1-50%                                                                               anti-                                           10-ol                          yeast                                           Flavor     2.0%         1-5%   flavor                                          Ethyl      48.0%       45-98.9%                                                                               diluent                                         Alcohol   100.0%                                                          ______________________________________                                    

B. DENTIFRICE

1. Liquid

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Liquid soap                                                                                5.0%         2-10%    surfac-                                      concentrate                       tant                                         Saccharin   0.2%         0.1-1.0% flavor                                       Clove Oil   1.0%         0.5-3.0% flavor                                       Cinnamon Oil                                                                               0.5%         0.5-3.0% flavor                                       Peppermint Oil                                                                             0.5%         0.5-3.0% flavor                                       Ethyl Alcohol                                                                              42.6%       29.5-95.3%                                                                               diluent                                      Color       0.2%         0.1-0.5% color                                        Limonen-10-ol                                                                              50.0%        1-50%    bacteri-                                                100.0%                 cide                                         ______________________________________                                    

2. Gel

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Sodium mono-                                                                               0.8%        0.5-1.5%  anti-                                        fluorophosphate                   plaque                                       Limonen-10-ol                                                                              50.0%       1-50%     bacteri-                                                                       cide                                         Hydrated silica                                                                            10.0%       8-15%     abrasive                                     xerogel                                                                        Hydrated    8.5%        5-10%     binder                                       thickening silica                                                              Sorbitol 70%                                                                               18.8%       5-73.3%   humectant                                    solution                                                                       Polyethylene                                                                               5.0%        3-7%      bodying                                      glycol 32                         agent                                        Sodium lauryl                                                                              1.5%        1-2%      surfac-                                      sulfate                           tant                                         Carboxymethyl                                                                              1.0%        0.5-2%    binder                                       cellulose gum                                                                  S D alcohol                                                                                1.0%        0.5-2%    stabi-                                                                         lizer                                        Flavor      3.0%        2-4%      flavor                                       Saccharin   0.2%        0.1-0.5%  flavor                                       F D & C Green                                                                              0.1%        0.1-0.5%  color                                        #3                                                                             F D & C Yellow                                                                             0.1%        0.1-0.5%  color                                        #10        100.0%                                                              ______________________________________                                    

3.Paste

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Sodium mono-                                                                               0.8%         0.5-1.5% anti-                                        fluorophosphate                   plaque                                       Limonen-10-ol                                                                              50.0%        1-50%    bacteri-                                                                       cide                                         Dicalcium   22.0%       20.4-30%  abrasive                                     phosphate                                                                      dihydrate                                                                      Water       16.0%       11.1-69.5%                                                                               diluent                                      Glycerine   5.1%         4.5-12.5%                                                                               bodying                                                                        agent                                        Flavor      2.0%         2-3%     flavor                                       Sodium lauryl                                                                              1.5%         1-2%     surfac-                                      sulfate                           tant                                         Carboxymethyl                                                                              1.4%         0.5-2.0% binder                                       cellulose gum                                                                  Tetrasodium                                                                                1.0%         0.5-2.0% binder                                       pyrophosphate                                                                  Sodium saccharin                                                                           0.2%         0.1-0.5% flavor                                                  100.0%                                                              ______________________________________                                    

C. OINTMENTS & SUPPOSITORIES WITH AND WITHOUT HYDROCORTISONE

1. Ointment With Hydrocortisone

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              1.0%         0.1-15.0%                                                                               bacteri-                                                                       cide                                         Polyethylene                                                                               59.5%       48.5-59.7%                                                                               bodying                                      glycol 3350                       agent &                                                                        emulsi-                                                                        fier                                         Polyethylene                                                                               39.5%       31.5-39.7%                                                                               bodying                                      glycol 400                        agent &                                                                        emulsi-                                                                        fier                                         Hydrocortisone                                                                             1.0%         0.5-5.0% anti-                                                   100.0%                 inflam-                                                                        matory                                       ______________________________________                                    

2. Ointment Without Hydrocortisone

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE      ACTION                                      ______________________________________                                         Limonen-10-ol                                                                              1.0%         0.1-15.0% anti-                                                                          yeast                                       Polyethylene                                                                               59.5        51.0-59.95%                                                                               bodying                                     glycol 3350                        agent &                                                                        emulsi-                                                                        fier                                        Polyethylene                                                                               39.5%       34.0-39.95%                                                                               bodying                                     glycol 400                         agent &                                                100.0%                  emuls-                                                                         ifier                                       ______________________________________                                    

3. Suppository Without Hydrocortisone

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE      ACTION                                      ______________________________________                                         Limonen-10-ol                                                                              1.0%         0.1-15.0% fungi-                                                                         cide                                        Polyethylene                                                                               9.5%        51.0-59.95%                                                                               bodying                                     glycol 1000                        agent &                                                                        emulsi-                                                                        fier                                        Polyethylene                                                                               39.5%       34.0-39.95%                                                                               bodying                                     glycol 3350                        agent &                                                100.0%                  emulsi-                                                                        fier                                        ______________________________________                                    

4. Suppository With Hydrocortisone

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              1.0%         0.1-15.0%                                                                               anti-                                                                          yeast                                        Polyethylene                                                                               74.0%       60.0-75.2%                                                                               bodying                                      glycol 1000                       agent &                                                                        emulsi-                                                                        fier                                         Polyethylene                                                                               24.0%       20.0-24.2%                                                                               bodying                                      glycol 3350                       agent &                                                                        emulsi-                                                                        fier                                         Hydrocortisone                                                                             1.0%         0.5-5.0% anti-                                                   100.0%                 inflam-                                                                        matory                                       ______________________________________                                    

D. Creams Without Hydorcortisone

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              1.0%         0.1-15.0%                                                                               bacteri-                                                                       cide                                         Cetyl alcohol                                                                              15.0%       12.0-18.0%                                                                               thickener                                    Arlacel 165                                                                                5.0%         3.5-7.5% emulsi-                                                                        fier                                         Sorbitol 70%                                                                               5.0%         3.5-8.0% humectant                                    solution                                                                       Water       74.5%       51.5-80.9%                                                                               diluent                                                 100.0%                                                              ______________________________________                                    

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              1.0%         0.1-15.0%                                                                               anti-                                                                          yeast                                        Spermaceti wax                                                                             12.5%       10.0-15.0%                                                                               thickener                                    Sorbitan                                                                       monostearate                                                                   Polyethylene 20                                                                            10.0%        7.5-12.5%                                                                               emulsi-                                                                        fier                                         Sorbitan    6.0%         4.0-8.0% emulsi-                                      monostearate                      fier                                         Water       75.5%       49.5-78.4%                                                                               diluent                                                 100.0%                                                              ______________________________________                                    

E. CREAMS WITH HYDROCORTISONE

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              1.0%         0.1-15.0%                                                                               anti-                                                                          yeast                                        Cetyl alcohol                                                                              15.0%       12.0-18.0%                                                                               thickener                                    Arlacel 165**                                                                              5.0%         3.5-7.5% emuls-                                                                         ifier                                        Sorbitol 70%                                                                               5.0%         3.5-8.0% humectant                                    solution                                                                       Hydrocortisone                                                                             1.0%         0.5-5.0% anti-                                                                          inflam-                                                                        matory                                       Water       73.0%       46.5-80.4%                                                                               diluent                                                 100.0%                                                              ______________________________________                                          *Croda, Inc., 51 Madison Ave., New York, New York 10010                        **Glycerol monostearate and polyoxyethylene stearate ICI of America            (Formerly Atlas Chemical Industries), Wilmington, Delaware 19899         

F. TAMPONS

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-    8.0%         1-15%    bacteri-                                     10-ol (2cc) 2Gm                   cide                                         Tampon 23Gm                                                                                92.0%       85-99%    reservoir                                               100.0%                 for bac-                                                                       tericide                                     ______________________________________                                    

G. AERSOLS WITHOUT HYDROCORTISONE

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              5.0%         0.5-50%  fungi-                                                                         cide                                         Ethyl alcohol                                                                              95.0%       50-99.5%  diluent                                                 100.0%                                                              ______________________________________                                          Pressurized nitrogen propellant at 100-125 psig                          

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              10.0%        0.5-50.0%                                                                               anti-                                                                          yeast                                        Soybean Oil                                                                                90.0%       50.0-99.5%                                                                               diluent                                                 100.0%                                                              ______________________________________                                          Pressurized nitrogen propellant at 100-125 psig                          

H. AERSOL WITH HYDROCORTISONE

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              10.0%        0.5-50.0%                                                                               bacteri-                                                                       cide                                         Soybean oil                                                                                98.0%       45-99.0%  diluent                                      Hydrocortisone                                                                             1.0%         0.5-5.0% anti-                                                   100.0%                 inflam-                                                                        matory                                       ______________________________________                                          Pressurized nitrogen propellant at 100-125 psig                          

I. OIL IN WATER EMULSION

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         (1) Limonen-    0.1%         0.1-50%                                                                               fungi-                                         10-ol                           cide                                       (2) Corn oil    10.0%       10-15%  oil                                        (2) Arlacel 40**                                                                               2.0%         1-3%   emulsi-                                                                        fier                                       (2) Tween 40    3.0%         2-4%   emulsi-                                                                        fier                                       (3) Water       84.9%       28-86.9%                                                                               diluent                                                   100.0%                                                          ______________________________________                                          Heat (2) to 70° C. Heat (3) to 72° C. Add (3) to (2) with        continuous agitation. When (3) and (2) cool to 40° C., add (1) wit      continuous agitation until room temperature is reached.                  

J. OIL IN WATER EMULSION WITH SOAP (FUNGICIDAL OR BACTERICIDAL SOAP)

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         (1) Limonen-    1.0%         0.1-25%                                                                               bacteri-                                       10-ol                           cide                                       (2) Corn oil    30.0%       20.0-40.0%                                                                             oil                                        (2) Arlacel 40**                                                                               2.0%         1.0-3.0%                                                                              emulsi-                                                                        fier                                       (2) Tween 40    3.0%         2.0-4.0%                                                                              emulsi-                                                                        fier                                       (2) Liquid soap                                                                                3.5%         2.5-5.0%                                                                              surfac-                                        concentrate                     tant                                       (3) Water       60.5%       23-74.4%                                                                               diluent                                                   100.0%                                                          ______________________________________                                          Heat (2) to 70° C. Heat (3) to 72° C. Add (3) to (2) with        continuous agitation. When (3) and (2) cool to 40° C., add (1) wit      continuous agitation until room temperature is reached.                  

K. WATER IN OIL EMULSION

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         (1) Limonen-    1.0%         0.1-25%                                                                               anti-                                          10-ol                           yeast                                      (2) Arlacel 186**                                                                              3.0%         2.0-4.0%                                                                              emulsi-                                                                        fier                                       (2) Soybean oil                                                                                15.0%       10.0-25.0%                                                                             oil                                        (2) Ceresin wax                                                                                0.5%         0.3-0.6%                                                                              thickener                                  (2) Beeswax     0.5%         0.3-0.6%                                                                              thickener                                  (2) Tween 80    0.5%         0.3-0.6%                                                                              emulsi-                                                                        fier                                       (3) Water       79.5%       44.2-87.0%                                                                             diluent                                                   100.0%                                                          ______________________________________                                          Heat (2) to 70° C. Heat (3) to 72° C. Add (3) to (2) with        continuous agitation. When (3) and (2) cool to 40° C., add (1) wit      continuous agitation until room temperature is reached.                  

L. PAINT

1. Enamel

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              1.00%        1-10%    bacteri-                                                                       cide                                         Titanium    14.91%      12-16%    pigment                                      dioxide                                                                        Calcium     29.83%      25-35%    pigment                                      carbonate                                                                      Silicate    4.81%        3-6%     pigment                                      Soya alkyd  25.72%      22-28%    pigment                                      resin                             (binder)                                     Mineral     23.73%       5-37%    solvent                                      spirits    100.00%                (thinner)                                    ______________________________________                                    

2. Latex

    ______________________________________                                         CHEMICAL   % OF TOTAL   RANGE     ACTION                                       ______________________________________                                         Limonen-10-ol                                                                              1.0%         1-10%    fungi-                                                                         cide                                         Titanium    10.76%       8-12%    pigment                                      dioxide                                                                        Silicate    12.91%      10-16%    pigment                                      Calcium     20.91%      15-25%    pigment                                      carbonate                                                                      Vinyl acrylic                                                                              12.22%      10-16%    vehicle                                      resin solids                      (binder)                                     Glycol      8.30%        6-10%    solvent                                                                        (thinner)                                    Water       34.00%      12-50%    solvent                                                 100.00%                (thinner)                                    ______________________________________                                    

While the invention has been described with respect to certain specific embodiments, it will be appreciated that many modifications and changes may be made by those skilled in the art without departing from the spirit of the invention. It is intended, therefore, by the appended claims to cover all such modifications and changes as fall within the true spirit and scope of the invention. 

What is claimed as new and what it is desired to secure by Letters Patent of the United States is:
 1. A method of killing bacteria, fungi or yeast comprising treating bacteria, fungi or yeast in their habitat with lethal concentrations of limonen-10-ol.
 2. The method of claim 1 wherein said bacteria are selected from a group consisting of Staphylococcus, Enterobacteriaceae, and Streptococcus.
 3. The method of claim 1 wherein said fungi are selected from a group consisting of Aureoblasidium and Cladosporium.
 4. A method of claim 1 for killing yeast wherein said yeast is Candida albicans. 